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The Zinc Nutritional Immunity of Epinephelus coioides Contributes to the Importance of znuC During Pseudomonas plecoglossicida Infection.

ABSTRACT: Previously, the dual RNA-seq was carried out in a Pseudomonas plecoglossicida- Epinephelus coioides infection model to investigate the dynamics of pathogen-host interplay in vivo. ZnuC, a member of ZnuCBA Zn importer, was found transcriptionally up-regulated during infection. Thus, this study aimed to assess its role during the trade-off for Zn between host and P. plecoglossicida. ICP-MS analysis and fluorescent staining showed that Zn was withheld from serum and accumulated in the spleen, with increased Zn uptake in the Golgi apparatus of macrophages after infection. Additionally, growth assay, macrophage infection and animal infection after gene knockout / silencing revealed that znuC was necessary for growth in Zn-limiting conditions, colonization, intracellular viability, immune escape and virulence of P. plecoglossicida. Further analysis with dual RNA-seq revealed associations of host's Zn nutritional immunity genes with bacterial Zn assimilation genes. IL6 and ZIP4 played key roles in this network, and markedly affected znuB expression, intracellular viability and immune escape, as revealed by gene silencing. Moreover, EMSA and GFP reporter gene analysis showed that Fur sensed changes in Fe concentration to regulate znuCBA in P. plecoglossicida. Jointly, these findings suggest a trade-off for Zn between host and P. plecoglossicida, while ZnuC is important for P. plecoglossicida Zn acquisition.

SUBMITTER: Huang L

PROVIDER: S-EPMC8129501 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Purpose: Next-generation sequencing (NGS) has revolutionized analysis of the immune response to V. alginolyticus in Epinephelus coioides larvae, we used high-throughput deep sequencing technology to study the effect of infection on gene expression. Methods: Epinephelus coioides larvae (30-days-old) were injected with V. alginolyticus , and fish were sacrificed at 24 h after infection. Ten wild-type Epinephelus coioides larvae were used to build an EST library for transcriptome analysis. The library products were prepared for sequencing analysis using an Illumina HiSeq™ 2000. Raw data were saved as fastq files. The following were removed: reads with adaptors, reads in which over 10% bases were unknown, and low quality reads (i.e., the percentage of bases of quality value ≤ 5 exceeds 50% in the read). Clean reads were mapped to reference sequences using SOAP aligner/soap2 . The randomness of RNA fragmentation was used to construct the library, and the numbers of reads mapped to the reference sequence were calculated. The RPKM method (Reads Per kb per Million reads) was used to calculate gene expression level, and differentially expressed genes (DEGs) Results: A total of 114,851,002 reads were assembled, consisting of 9,687,355,560 nucleotides; these were further assembled into 209,082 contigs with a mean length of 372 bp. Gene ontology (GO) analysis of the transcriptome revealed 12 cellular component subcategories, 16 molecular function subcategories, and 42 biological process subcategories (P value < 0.05). A total of 32664 Epinephelus coioides genes were mapped to the Kyoto Encyclopedia of Genes and Genomes (KEGG); 1504 differentially expressed genes (DEGs) were subsequently identified, in 12 categories (P value < 0.05). Vibrio infection affected the expression of genes involved in complementation, coagulation cascades, pathogen (Staphylococcus aureus) infection, phagosome activity, antigen processing, and the antigen presentation pathway. Conclusions: We conclude that the complement pathway of innate immunity and the hepicidin antimicrobial peptide may play important roles in the defense of Epinephelus coioides larvae against V. alginolyticus, and the immune response may activate at 4h after bacterial infection. These results implicate the complement pathway signal pathway in immunity during V. alginolyticus infection at early developmental stages, enhancing our understanding of the mechanisms underlying the immune response to Vibrio infection in Epinephelus coioides.

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The Zinc Nutritional Immunity of Epinephelus coioides Contributes to the Importance of znuC During Pseudomonas plecoglossicida Infection.

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